DESCRIPTION: The opportunistic pathogen Pseudomonas aeruginosa is capable of responding rapidly to changes in the environment. During an infection, P. aeruginosa is capable of selectively activating or repressing classes of genes which are important for their survival in the human host. Using an IVET selection scheme adapted from techniques used to study Salmonella and Vibrio pathogenesis, several genes were identified and shown to be transcriptionally activated when P. aeruginosa was grown in vivo. These same genes were poorly expressed when the P. aeruginosa was grown under standard laboratory conditions. One such gene, ppkA, encodes a homologue of eukaryotic serine/threonine protein kinases. Subsequent studies identified a second similar gene designated ppkB. These observations suggest that P. aeruginosa may possess a unique serine/threonine kinase-based signaling network. This proposal will test the hypothesis that the ppkA/ppkB gene products play a role in P. aeruginosa pathogenesis by signaling and promoting the expression of important virulence factors. Isogenic wild-type, ppkA, and ppkB deficient P. aeruginosa strains will be tested for virulence in several animal models of infections. Biochemical and genetic characterization of PpkA will be undertaken to determine the site(s) of phosphorylation and to identify additional proteins which interact with PpkA. Finally genes which may be targets of PpkA or PpkB proteins will be identified and their products characterized to determine if they play a role in P. aeruginosa pathogenesis. These studies should yield information about this novel serine/threonine signaling pathway in P. aeruginosa and may be a model for similar regulatory pathways in other bacteria.